Studies in the field of cell cycle regulation have identified many important cell cycle regulators and provided a wealth of knowledge about how cell proliferation is controlled in various settings. The kinase inhibitor p27Kipl inhibits the kinase activity of various cyclin-dependent kinases to restrain cell proliferation. Targeted inactivation of p27 in the mouse results in proportionally enlarged animals with multi-organomegaly. The livers of these animals contain about 25% more apparently normal hepatocytes but do not have increased incidence of hepatocellular carcinoma. We reasoned that this property of p27 in hepatocytes may be used to benefit hepatocyte-based therapies. We have recently reported that hepatocytes isolated from p27 knockout mice demonstrated improved proliferation ability in a mouse liver failure transplantation model, providing initial experimental support for our hypothesis that manipulation of p27 could be a useful approach to improving the efficiency of hepatocyte transplantation. The objective of this application is to gain a better understanding of p27 in hepatocytes with next-level p27 targeting strategies, which will then enable us to better evaluate the potentials of targeting p27 in human hepatocyte transplantation. In Specific Aim 1, we will generate hepatocyte-specific, inducible p23 knockout mice to study the role of p23 in hepatocytes at various stages in the liver. We will then use hepatocytes isolated from these mice to determine the functional role and mechanisms of p27 in hepatocyte proliferation in vitro (Specific Aim 2) and after transplantation (Specific Aim 3). In Specific Aim 4, we will determine the role of p27 in hepatocellular carcinogenesis in two mouse liver tumor models that involve extensive hepatocyte proliferation prior to oncogenic transformation. Successful completion of these studies will provide a solid knowledge of p27 in hepatocytes. This knowledge will form the foundation by which hepatocyte proliferation is controlled, which is essential for finding rationale ways to manipulate hepatocyte proliferation to benefit the treatment of liver diseases and liver gene therapy.